The study of human cells defective in repairing damaged DNA was extended, with the rationale that DNA-repair deficient cells are more susceptible to the adverse effects of carcinogens (cell killing, mutagenesis, sister chromatid exchange, and malignant transformation) than their repair-proficient counterparts. A group of human tumor (17) and SV40-transformed (7) strains deficient in the repair of 0-six-methylguanine (0-6-MeG, a modified DNA base made by certain methylating agents) was identified earlier in this project. Such strains were called MER (-). Like SV40, Rous sarcoma virus was found to produce MER (-) strains. After treatment to produce 0-6-MeG, MER (-) strains, while able to relax the supercoiling of their DNA as well as MER (+) strains, were unable to restore supercoiling to their DNA, whereas MER (+) strains could. One MER (-) strain was found to be unable to relax DNA supercoiling after such treatment, leading to the idea that the relaxation process is enzymatic. Ethidium bromide titration techniques were employed as a further probe into the nature of the initial relaxation step characteristic of cells proficient and deficient in the repair of both ultraviolet- or methyl-damaged DNA. A activity that removes the methyl group from 0-six-methylguanine contained in DNA was detected in MER (+) but not MER (-) lines.